This invention relates to new amidine derivatives, processes for their preparation, compositions containing them and their use in therapy.
Nitric oxide is produced in mammalian cells from L-arginine by the action of specific nitric oxide synthases (NOSs). These enzymes fall into two distinct classesxe2x80x94constitutive NOS (cNOS) and inducible NOS (iNOS). At the present time, two constitutive NOSs and one inducible NOS have been identified. Of the constitutive NOSs, an endothelial enzyme (ecNOS) is involved with smooth muscle relaxation and the regulation of blood pressure and blood flow, whereas the neuronal enzyme (ncNOS) serves as a neurotransmitter and appears to be involved in the regulation of various biological functions such as cerebral ischaemia. Inducible NOS has been implicated in the pathogenesis of inflammatory diseases. Specific regulation of these enzymes should therefore offer considerable potential in the treatment of a wide variety of disease states.
Compounds of various structures have been described as inhibitors of NOS and their use in therapy has been claimed. See, for example, WO 95/09619 (The Wellcome Foundation) and WO 95/11231 (G. D. Searle). The applicant has previously disclosed in WO 95/05363 and WO 96/01817 amidine derivatives which are NOS inhibitors which display some selectivity for inhibition of the neuronal enzyme, ncNOS.
We now disclose a group of amidines that are within the generic scope of WO 96/01817, but which are not specifically exemplified in WO 96/01817. These compounds display surprisingly advantageous properties and are the subject of the present application.
According to the invention we provide a compound of formula (I) 
wherein:
R1 represents a 2-thienyl or 3-thienyl ring;
and R2 represents hydrogen or C 1 to 4 alkyl;
and optical isomers and racemates thereof and pharmaceutically acceptable salts thereof.
Preferably R1 represents 2-thienyl.
Preferably R2 represents hydrogen, methyl or 2-propyl.
Particularly preferred compounds of the invention include:
N-(4-methyl-2,3,4,5-tetrahydro-1,4-benzoxazepin-7-yl)-2-thiophenecarboximidamide;
N-(4-ethyl-2,3,4,5-tetrahydro-1,4-benzoxazepin-7-yl))-2-thiophenecarboximidamide;
N-(4-propyl-2,3,4,5-tetrahydro-1,4-benzoxazepin-7-yl)-2-thiophenecarboximidamide;
N-(4-isopropyl-2,3,4,5-tetrahydro-1,4-benzoxazepin-7-yl)-2-thiophenecarboximidamide;
N-(2,3,4,5-tetrahydro-1,4-benzoxazepin-7-yl)-2-thiophenecarboximidamide;
N-(2,3,4,5-tetrahydro-1,4-benzoxazepin-7-yl)-3-thiophenecarboximidamide;
N-(4-methyl-2,3,4,5-tetrahydro-1,4-benzoxazepin-7-yl)-3-thiophenecarboximidamide; and pharmaceutically acceptable salts thereof.
More especially preferred compounds of the invention include:
N-(4-methyl-2,3,4,5-tetrahydro-1,4-benzoxazepin-7-yl)-2-thiophenecarboximidamide;
N-(4-isopropyl-2,3,4,5-tetrahydro-1,4-benzoxazepin-7-yl)-2-thiophenecarboximidamide;
N-(2,3,4,5-tetrahydro-1,4-benzoxazepin-7-yl)-2-thiophenecarboximidamide; and pharmaceutically acceptable salts thereof.
Unless otherwise indicated, the term xe2x80x9cC 1 to 4 alkylxe2x80x9d referred to herein denotes a straight or branched chain alkyl group having from 1 to 4 carbon atoms. Examples of such groups include methyl, ethyl, n-propyl, i-propyl, n-butyl, i-butyl and t-butyl.
The present invention includes compounds of formula (I) in the form of salts, in particular acid addition salts. Suitable salts include those formed with both organic and inorganic acids. Such acid addition salts will normally be pharmaceutically acceptable although salts of non-pharmaceutically acceptable acids may be of utility in the preparation and purification of the compound in question. Thus, preferred salts include those formed from hydrochloric, hydrobromic, sulphuric, phosphoric, citric, tartaric, lactic, pyruvic, acetic, succinic, fumaric, maleic, methanesulphonic and benzenesulphonic acids.
According to the invention, we further provide a process for the preparation of compounds of formula (I), and optical isomers and racemates thereof and pharmaceutically acceptable salts thereof, which comprises:
(a) preparing a compound of formula (I) by reacting a corresponding compound of formula (II) 
wherein R2 is as defined above,
with a compound of formula (III) or an acid addition salt thereof 
wherein R1 is as defined above and L is a leaving group;
(b) preparing a compound of formula (I) by reacting a corresponding compound of formula (IV) 
wherein R2 is as defined above and HA is an acid,
with a compound of formula (V)
R1xe2x80x94xe2x89xa1Nxe2x80x83xe2x80x83(V)
wherein R1 is as defined above;
(c) preparing a compound of formula (I) in which R2 represents C 1 to 4 alkyl by reacting a corresponding compound of formula (I) in which R2 represents hydrogen with a compound of formula (VI)
R3xe2x80x94Lxe2x80x83xe2x80x83(VI)
wherein R3 represents C 1 to 4 alkyl and L is a leaving group; or
(d) preparing a compound of formula (I) in which R2 represents methyl by reacting a corresponding compound of formula (I) in which R2 represents hydrogen with formaldehyde and formic acid;
and where desired or necessary converting the resultant compound of formula (I), or another salt thereof, into a pharmaceutically acceptable salt thereof, or vice versa, and where desired converting the resultant compound of formula (I) into an optical isomer thereof
In process (a), the reaction will take place on stirring a mixture of the reactants in a suitable solvent, for example, N-methyl-2-pyrrolidinone or a lower alkanol such as ethanol, isopropanol or tertiary butanol, at a temperature between room temperature and the reflux temperature of the solvent. The reaction time will depend inter alia on the solvent and the nature of the leaving group, and may be up to 48 hours; however it will typically be from 1 to 24 hours. Suitable leaving groups that L may represent include thioalkyl, sulphonyl, trifluoromethyl sulphonyl, halide, alkyl alcohols, aryl alcohols and tosyl groups; others are recited in xe2x80x98Advanced Organic Chemistryxe2x80x99, J. March (1 985) 3rd Edition, on page 315 and are well known in the art.
In process (b), the reaction is preferably performed by refluxing a mixture of the two compounds for several hours in the presence of a suitable solvent whereby the reaction temperature is high enough so that condensation takes place readily, but not sufficiently high to decompose the amidine formed. The reaction temperature can vary from room temperature to about 250xc2x0 C., although it is preferable to perform the reaction at temperatures from about 100xc2x0 C. to 200 xc2x0 C. We find that o-dichlorobenzene is a particularly suitable solvent. We also find that it is often useful to add 4-dimethylaminopyridine as a catalyst. On cooling, two layers form, the solvent may be decanted, and the reaction worked up by addition of aqueous base. Alternatively, where the reactants are soluble in the solvent, the solvent may be evaporated off under vacuum and the reaction mixture worked up by addition of water. The acid HA may be an organic or inorganic acid, for instance, hydrochloric, hydrobromic, hydroiodic, sulphuric, nitric, phosphoric, acetic, lactic, succinic, fumaric, malic, maleic, tartaric, citric, benzoic or methanesulphonic acid. We prefer that HA is a hydrohalic acid.
In process (c) the reaction will take place under standard conditions, for example by reacting the two compounds in an inert solvent such as DMF under basic conditions at a suitable temperature, typically room temperature, for a period of up to 72 hours or until the reaction is complete. We have frequently found it desirable to treat the amine with NaH before reacting with the compound of formula (VI). Suitable leaving groups L are mentioned above. We prefer that L represents halide, particularly bromide.
In process (d), the reaction will typically take place on refluxing the reaction mixture for up to 4 hours or until reaction is complete.
Salts of compounds of formula (I) may be formed by reacting the free base or a salt, enantiomer, tautomer or protected derivative thereof, with one or more equivalents of the appropriate acid. The reaction may be carried out in a solvent or medium in which the salt is insoluble, or in a solvent in which the salt is soluble followed by subsequent removal of the solvent in vacuo or by freeze drying. Suitable solvents include, for example, water, dioxan, ethanol, isopropanol, tetrahydrofuran or diethyl ether, or mixtures thereof. The reaction may be a metathetical process or it may be carried out on an ion exchange resin.
The compounds of formula (II) may be prepared by reduction of a corresponding compound of formula (VII) 
wherein R2 is as defined above.
The reduction reaction may be performed under a number of conditions, for example those described in J. March xe2x80x9cAdvanced Organic Chemistryxe2x80x9d on pages 1103-1104. These include catalytic hydrogenation, use of Zn, Sn or Fe metal, AlH3xe2x80x94AlCl3, sulphides and others. We prefer to perform the reaction by hydrogenation at atmospheric pressure in the presence of a palladium and carbon catalyst until reaction is complete, typically for 3 to 6 hours, or by reduction using zinc metal in acetic acid and methanol.
Compounds of formula (VII) may be prepared by cyclising a compound of formula (VIII) 
wherein R2 is as defined above and L is a leaving group, preferably fluoro;
or by cyclising a compound of formula (IX) 
wherein R2 is as defined above and L is a leaving group.
Compounds of formula (VIII) may be prepared by reaction of a compound of formula (X) 
wherein L is a leaving group, preferably fluoro,
with a compound of formula (XI) 
wherein R2 is as defined above, by the process of reductive amination.
Other syntheses of compounds of formula (VIII) and (IX) will be readily apparent to one skilled in the art. Compounds of formula (VIII) or (IX) may cyclise directly to a compound of formula (VII) without the need for prior isolation. The cyclisation reactions may also take place on removal of protecting groups. In the above reactions it may be desirable to render the nucleophilic group xe2x80x94OH in compounds of formula (VIII) and (IX) more reactive by treatment with base.
Compounds of formula (VII) may also be prepared by nitration of a compound of formula (XII) 
wherein R2 is as defined above.
The nitration reaction will take place under conditions well known to a person skilled in the art, for example, on treatment with nitric acid and sulphuric acid or potassium nitrate and sulphuric acid, optionally in an inert organic solvent.
It may also be convenient to prepare compounds of formula (VII) by nitration of a carbonyl or dicarbonyl derivative of a compound of formula (XII); which nitrated carbonyl or dicarbonyl derivative may be reduced to the desired compound of formula (VII) using, for example, diborane.
Compounds of formula (VII) and (XII), as well as certain carbonyl and dicarbonyl derivatives of compounds of formula (XII) just mentioned may also be prepared by one of the numerous methods for preparation of bicyclic heterocyclic compounds.
Compounds of formula (XII) in which R2 represents hydrogen may also be prepared by a synthesis based on ring expansion to convert a cyclic ketone into a cyclic amide (Grunewald and Dahanukar, J. Heterocyclic Chem., 1994, 31, 1609-1617).
Thus, a compound of formula (XIII) 
may be converted into a compound of formula (XIV) 
on treatment with sodium azide in acid. Further details of the reaction conditions may be obtained by reference to the above mentioned Grunewald and Dahanukar paper.
It will be apparent to a person skilled in the art that the compounds of formula (XIV) may also desirably be prepared in nitrated form. Nitration may be achieved by treatment of the non-nitrated analogue with nitric acid and sulphuric acid or potassium nitrate and sulphuric acid under standard conditions.
Intermediate compounds may be prepared as such or in protected form. In particular amine and hydroxyl groups may be protected. Suitable protecting groups are described in the standard text xe2x80x9cProtective Groups in Organic Synthesisxe2x80x9d, 2nd Edition (1991) by Greene and Wuts. Amine-protecting groups which may be mentioned include alkyloxycarbonyl such as t-butyloxycarbonyl, phenylalkyloxycarbonyl such as benzyloxycarbonyl, or trifluoroacetate. Deprotection will normally take place on treatment with aqueous base or aqueous acid.
Compounds of formula (VII), (VIII), (IX), (XI) and (XII) in which R2 represents C 1 to 4 alkyl may also be prepared by alkylating the corresponding compound in which R2 represents hydrogen following process (c) above.
Compounds of formula (IV) may be prepared by analogous processes to those described for the preparation of compounds of formula (II). Compounds of formula (IV) may be converted into corresponding compounds of formula (II) by treatment with a base. Compounds of formula (II) may be converted into corresponding compounds of formula (IV) by treatment with a protic acid HA, for example, one of those listed above.
Compounds of formula (III) are either known or may be prepared by known methods. For example, compounds of formula (III) in which L represents thioalkyl may be prepared by treatment of the corresponding thioamide of formula (XV) 
wherein R1 is as defined above, with an alkylhalide under conditions well known to a person skilled in the art.
Alternatively, the acid addition salts of compounds of formula (III) wherein L is thioalkyl may be prepared by reaction of a nitrile of formula (V) with an alkyl thiol and acid, for example hydrochloric acid, in a solvent such as dichloromethane or diethyl ether.
Compounds of formula (V), (VI), (X), (XI), (XIII), (XIV) and (XV) are either known or may be prepared by conventional methods known per se.
It will be apparent to a person skilled in the art that it may be desirable to protect an amine or other reactive group in an intermediate compound using a protecting group as described in the standard text xe2x80x9cProtective Groups in Organic Synthesisxe2x80x9d, 2nd Edition (1991) by Greene and Wuts. Suitable amine-protecting groups are mentioned above.
The compounds of the invention and intermediates may be isolated from their reaction mixtures, and if necessary further purified, by using standard techniques.
The compounds of formula (I) may exist in tautomeric, enantiomeric or diastereoisomeric forms, all of which are included within the scope of the invention. The various optical isomers may be isolated by separation of a racemic mixture of the compounds using conventional techniques, for example, fractional crystallisation or HPLC. Alternatively, the individual enantiomers may be made by reaction of the appropriate optically active starting materials under reaction conditions which will not cause racemisation.
Intermediate compounds may also exist in enantiomeric forms and may be used as purified enantiomers, diastereomers, racemates or mixtures.
The compounds of general formula (I) possess useful nitric oxide synthase inhibiting activity, and in particular, they exhibit good selectivity for inhibition of the neuronal isoform of nitric oxide synthase. They are thus useful in the treatment or prophylaxis of human diseases or conditions in which the synthesis or oversynthesis of nitric oxide by nitric oxide synthase forms a contributory part. Examples of such diseases or conditions include hypoxia, such as in cases of cardiac arrest, stroke and neonatal hypoxia, neurodegenerative conditions including nerve degeneration and/or nerve necrosis in disorders such as ischaemia, hypoxia, hypoglycemia, epilepsy, and in external wounds (such as spinal cord and head injury), hyperbaric oxygen convulsions and toxicity, dementia, for example, pre-senile dementia, Alzheimer""s disease and AIDS-related dementia, Sydenham""s chorea, Parkinson""s disease, Huntington""s disease, Amyotrophic Lateral Sclerosis, Korsakoffs disease, imbecility relating to a cerebral vessel disorder, sleeping disorders, schizophrenia, anxiety, depression, seasonal affective disorder, jet-lag, depression or other symptoms associated with Premenstrual Syndrome (PMS), anxiety and septic shock. The compounds of formula (I) are also useful in the treatment and alleviation of acute or persistent inflammatory or neuropathic pain, or pain of central origin, and in the treatment or prophylaxis of inflammation. Compounds of formula (I) may also be expected to show activity in the prevention and reversal of tolerance to opiates and diazepines, treatment of drug addiction and treatment of migraine and other vascular headaches. The compounds of the present invention may also show useful immunosuppressive activity, and be useful in the treatment of gastrointestinal motility disorders, and in the induction of labour. The compounds may also be useful in the treatment of cancers that express nitric oxide synthase.
Compounds of formula (I) are predicted to be particularly useful in the treatment or prophylaxis of hypoxia or stroke or ischaemia or neurodegenerative conditions or schizophrenia or migraine or for the prevention and reversal of tolerance to opiates and diazepines or for the treatment of drug addiction or for the treatment of pain and especially in the treatment or prophylaxis of hypoxia or stroke or ischaemia or neurodegenerative disorders or schizophrenia or pain. We are particularly interested in conditions selected from the group consisting of hypoxia, ischaemia, stroke, pain, schizophrenia, Parkinson""s disease, Huntington""s disease and Amyotrophic Lateral Sclerosis.
For the treatment of pain, the compounds of formula (I) are expected to be particularly useful either alone, or in combination with other agents such as opiates, particularly morphine.
For the treatment of Parkinson""s disease, the compounds of formula (I) are expected to be particularly useful either alone, or in combination with other agents such as L-Dopa.
Prophylaxis is expected to be particularly relevant to the treatment of persons who have suffered a previous episode of, or are otherwise considered to be at increased risk of, the disease or condition in question. Persons at risk of developing a particular disease or condition generally include those having a family history of the disease or condition, or those who have been identified by genetic testing or screening to be particularly susceptible to developing the disease or condition.
Thus according to a further aspect of the invention we provide a compound of formula (I), or an optical isomer or racemate thereof or a pharmaceutically acceptable salt thereof, for use as a medicament.
According to another feature of the invention we provide the use of a compound of formula (I) or an optical isomer or racemate thereof or a pharmaceutically acceptable salt thereof, in the manufacture of a medicament for the treatment or prophylaxis of the aforementioned diseases or conditions; and a method of treatment or prophylaxis of one of the aforementioned diseases or conditions which comprises administering a therapeutically effective amount of a compound of formula (I), or an optical isomer or racemate thereof or a pharmaceutically acceptable salt thereof, to a person suffering from or susceptible to such a disease or condition.
For the above mentioned therapeutic indications, the dosage administered will, of course, vary with the compound employed, the mode of administration and the treatment desired. However, in general, satisfactory results are obtained when the compounds are administered to a human at a daily dosage of between 0.5 mg and 2000 mg (measured as the active ingredient) per day, particularly at a daily dosage of between 2 mg and 500 mg.
The compounds of formula (I), and optical isomers and racemates thereof and pharmaceutically acceptable salts thereof, may be used on their own, or in the form of appropriate medicinal formulations. Administration may be by, but is not limited to, enteral (including oral, sublingual or rectal), intranasal, or topical or other parenteral routes. Conventional procedures for the selection and preparation of suitable pharmaceutical formulations are described in, for example, xe2x80x9cPharmaceuticalsxe2x80x94The Science of Dosage Form Designsxe2x80x9d, M. E. Aulton, Churchill Livingstone, 1988.
According to the invention, there is provided a pharmaceutical formulation comprising preferably less than 95% by weight and more preferably less than 50% by weight of a compound of formula (I), or an optical isomer or racemate thereof or a pharmaceutically acceptable salt thereof, in admixture with a pharmaceutically acceptable diluent or carrier. The formulation may optionally also contain a second pharmacologically active ingredient such as L-Dopa, or an opiate analgesic such as morphine.
We also provide a method of preparation of such a pharmaceutical formulation which comprises mixing the ingredients.
Examples of such diluents and carriers are: for tablets and dragees: lactose, starch, talc, stearic acid; for capsules: tartaric acid or lactose; for injectable solutions: water, alcohols, glycerin, vegetable oils; for suppositories: natural or hardened oils or waxes.
Compositions in a form suitable for oral, that is oesophageal, administration include: tablets, capsules and dragees; sustained release compositions include those in which the active ingredient is bound to an ion exchange resin which is optionally coated with a diffusion barrier to modify the release properties of the resin.
The enzyme nitric oxide synthase has a number of isoforms and compounds of formula (I), and optical isomers and racemates thereof and pharmaceutically acceptable salts thereof, may be screened for nitric oxide synthase inhibiting activity by following procedures based on those of Bredt and Snyder in Proc. Natl. Acad. Sci., 1990, 87, 682-685. Nitric oxide synthase converts 3H-L-arginine into 3H-L-citrulline which can be separated by cation exchange chromatography and quantified by scintillation counting.
Screen for Neuronal Nitric Oxide Synthase Inhibiting Activity
The enzyme is isolated from rat hippocampus or cerebellum. The cerebellum or hippocampus of a male Sprague-Dawley rat (250-275 g) is removed following CO2 anaesthesia of the animal and decapitation. Cerebellar or hippocampal supernatant is prepared by homogenisation in 50 mM Tris-HCl with 1 mM EDTA buffer (pH 7.2 at 25xc2x0 C.) and centifugation for 15 minutes at 20,000 g. Residual L-arginine is removed from the supernatant by chromatography through Dowex AG-50W-X8 sodium form and 25 hydrogen form columns successively, and further centrifugation at 1000 g for 30 seconds.
For the assay, 25 xcexcl of the final supernatant is added to each of 96 wells (of a 96 well filter plate) containing either 25 xcexcl of an assay buffer (50 mM HEPES, 1 mM EDTA, 1.5 mM CaCl2, pH 7.4) or 25 xcexcl of test compound in the buffer at 22xc2x0 C. and 25 xcexcl of complete assay buffer (50 mM HEPES, 1 mM EDTA, 1.5 mM CaCl2, 1 mM DTT, 100 xcexcM NADPH, 10 xcexcg/ml calmodulin, pH 7.4). Following a 10 minute equilibration period, 25 xcexcl of an L-arginine solution (of concentration 18 xcexcM 1H-L-arginine, 96 nM 3H-L-arginine) is added to each well to initiate the reaction. The reaction is stopped after 10 minutes by addition of 200 xcexcl of a slurry of termination buffer (20 mM HEPES, 2 mM EDTA, pH 5.5) and Dowex AG-50W-X8 200-400 mesh.
Labelled L-citrulline is separated from labelled L-arginine by filtering each filter plate and 75 xcexcl of each terminated reaction is added to 3 ml of scintillation cocktail. The L-citrulline is then quantified by scintillation counting.
In a typical experiment using the cerebellar supernatant, basal activity is increased by 20,000 dpm/ml of sample above a reagent blank which has an activity of 7,000 dpm/ml. A reference standard, N-nitro-L-arginine, which gives 80% inhibition of nitric oxide synthase at a concentration of 1 xcexcM, is tested in the assay to verify the procedure.
Screen for Endothelial Nitric Oxide Synthase Inhibiting Activity
The enzyme is isolated from human umbilical vein endothelial cells (HUVECs) by a procedure based on that of Pollock et al in Proc. Natl. Acad. Sci., 1991, 88, 10480-10484. HUVECs were purchased from Clonetics Corp (San Diego, Calif., USA) and cultured to confluency. Cells can be maintained to passage 35-40 without significant loss of yield of nitric oxide synthase. When cells reach confluency, they are resuspended in Dulbecco""s phosphate buffered saline, centrifuged at 800 rpm for 10 minutes, and the cell pellet is then homogenised in ice-cold 50 mM Tris-HCl, 1 mM EDTA, 10% glycerol, 1 mM phenylmethylsulphonylfluoride, 2 xcexcM leupeptin at pH 4.2. Following centrifugation at 34,000 rpm for 60 minutes, the pellet is solubilised in the homogenisation buffer which also contains 20 mM CHAPS. After a 30 minute incubation on ice, the suspension is centrifuged at 34,000 rpm for 30 minutes. The resulting supernatant is stored at xe2x88x9280xc2x0 C. until use.
For the assay, 25 xcexcl of the final supernatant is added to each of 12 test tubes containing 25 xcexcl L-arginine solution (of concentration 12 xcexcM 1H-L-arginine, 64 nM 3H-L-arginine) and either 25 xcexcl of an assay buffer (50 mM HEPES, 1 mM EDTA, 1.5 mM CaCl2, pH 7.4) or 25 xcexcl of test compound in the buffer at 22xc2x0 C. To each test tube was added 25 xcexcl of complete assay buffer (50 mM HEPES, 1 mM EDTA, 1.5 mM CaCl2, 1 mM DTT, 100 xcexcM NADPH, 10 xcexcg/ml calmodulin, 12 xcexcM tetrahydrobiopterin, pH 7.4) to initiate the reaction and the reaction is stopped after 10 minutes by addition of 2 ml of a termination buffer (20 mM HEPES, 2 mM EDTA, pH 5.5).
Labelled L-citrulline is separated from labelled L-arginine by chromatography over a Dowex AG-50W-X8 200-400 mesh column. A 1 ml portion of each terminated reaction mixture is added to an individual I ml column and the eluant combined with that from two 1 ml distilled water washes and 16 ml of scintillation cocktail. The L-citrulline is then quantified by scintillation counting.
In a typical experiment, basal activity is increased by 5,000 dpm/ml of sample above a reagent blank which has an activity of 1500 dprn/ml. A reference standard, N-nitro-L-arginine, which gives 70-90% inhibition of nitric oxide synthetase at a concentration of 1 xcexcM, is tested in the assay to verify the procedure.
In the screens for nitric oxide synthase inhibition activity, compound activity is expressed as IC50 (the concentration of drug substance which gives 50% enzyme inhibition in the assay). IC50 values for test compounds were initially estimated from the inhibiting activity of 1, 10 and 100 xcexcM solutions of the compounds. Compounds that inhibited the enzyme by at least 50% at 10 xcexcM were re-tested using more appropriate concentrations so that an IC50 could be determined.
When tested in the above screens, the compounds of Examples 1 to 7 below showed lC50 values for inhibition of neuronal nitric oxide synthase of less than 10 xcexcM and good selectivity for inhibition of the neuronal isoform of the enzyme, indicating that they are predicted to show particularly useful therapeutic activity.
When compared with other compounds, the compounds of formula (I), and optical isomers and racemates thereof and pharmaceutically acceptable salts thereof, have the advantage that they may be less toxic, be more efficacious, be longer acting, have a broader range of activity, be more potent, be more selective for the neuronal isoform of nitric oxide synthase enzyme, produce fewer side effects, be more easily absorbed or have other useful pharmacological properties.
The invention is illustrated by the following examples: